We report direct evidence for the formation of a functional enzyme-enzyme-substrate complex between glyceraldehyde-3-PO4 dehydrogenase (GPDH) and phosphoglycerate kinase-1, 3 diphosphoglycerate (PGK-DPG) complex. Reduction by NADH proceeds exclusively via the E.E'-S complex. Interaction between the two enzymes is enhanced by DPG ligation. The affinity for cognate species of enzymes appears to be greater than between hybrid species. The ATPase driven translocation of cations by Na/K ATPase and by Ca ATPase can be studied by utilizing a chromophoric high energy phosphate substrate, Beta-furylacryloyl-phosphate (FAP). Transient studies with Ca ATPase are strikingly similar to results obtained with ATP as substrate. These results demonstrate the rapid for formation of a phosphoenzyme with concomitant enhancement of Ca binding and ion-translocation, followed by slower hydrolysis of the phosphoenzyme. In contrast, FAP hydrolysis by the N/K ATPase to follow a strict "Phosphatase" mechanism with no evident phosphoenzyme formation. The ATPase mechanism has been shown to be similar with the two enzymes. Evidence is presented for facile uncoupling of the phosphorylation and the phosphatase activities of the Na/K ATPase by detergent solubilization; a process which is reversed by nucleotide binding. The stable coupled structure of sarcoplasmicreticulum Ca ATPase makes the phosphorylation-phosphatase pathway obligatory, even with the pseudo substrate.